Diabetes mellitus is a huge health burden due to decreased quality of life and the escalating cost of treatment. In the United States alone, ~8% of the population is diabetic and nearly one-third of adults are now estimated to be at risk to develop the disease. Obesity, insulin resistance and metabolic abnormalities in liver, adipose, and muscle are important factors in disease. Yet, most of the gene loci recently found associated with type 2 diabetes encode proteins that enable insulin production from pancreatic cells. Nutrients and hormones regulate not only insulin secretion but also the capacity of cells to continue to produce insulin. In this proposal we focus on mechanisms of action of nutrients and agents that enhance insulin production in cells. The overarching goals are to elucidate mechanisms that can be manipulated to improve -cell function and overcome the physiological changes in cells that occur during prolonged hyperglycemia, contributing to inadequate insulin release. In aim 1 we will determine the functions of the taste receptor complex (T1R1/T1R3) in pancreatic cells. This dimeric G protein-coupled receptor (GPR) is involved in amino acid-sensing. This receptor, identified in gustatory neurons, binds to a variety of amino acids, but has not been studied on cells. We find that stable knockdown of T1R3 reduces insulin secretion, and insulin content, and causes events associated with autophagy. We will explore the bases for these phenotypic changes and we will also examine the underlying signaling mechanisms. In the second aim we will examine molecular mechanisms of action of small molecules that enhance beta-cell function. These molecules stimulate insulin production by cells, improve oral glucose tolerance of db/db mice, and restore insulin production by human islets in long term culture. We have identified a number of changes that take place in cells treated with these drugs, including epigenetic alterations, and changes in concentrations of key transcription factors. We will evaluate candidates that may mediate the actions of these drugs identified by drug-affinity chromatography. Finally, we will continue to investigate ERK1/2-dependent signaling events that control insulin gene transcription by focusing on mechanisms of recruitment of the coactivator p300. These studies should provide new understanding of how amino acids regulate insulin production, stability and release and will exploit a new pharmacological tool to identify mechanisms to enhance insulin production and insulin transcription in normal and failing islets.